Phosphorylated materials as contrast agents for use in magnetic resonance imaging of the gastrointestinal region

ABSTRACT

Methods of providing an image of the gastrointestinal region of a patient and diagnosing the presence of any tumorous tissue in that region using contrast media comprising a combination of at least one polyphosphorylated aliphatic or polyphosphorylated alicyclic compound and at least one paramagnetic ion, wherein the aliphatic and alicyclic compounds comprise at least five carbon atoms, is described. Also described are diagnostic kits for gastrointestinal imaging which include the subject contrast media.

CROSS-REFERENCE TO RELATED APPLICATIONS

This is a divisional of U.S. application Ser. No. 08/435,489, filed May5, 1995, now U.S. Pat. No. 5,525,326, which is a divisional of U.S.application Ser. No. 08/164,018, filed Feb. 17, 1994, now U.S. Pat. No.5,449,508, which is a divisional of U.S. application Ser. No.07/867,166, filed Apr. 10, 1992, now U.S. Pat No. 5,320,826, which is adivisional of U.S. application Ser. No. 07/649,437, filed Feb. 1, 1991,now U.S. Pat No. 5,143,716.

BACKGROUND OF THE INVENTION

Magnetic Resonance Imaging (MRI) is a relatively new diagnostic imagingtechnique which employs a magnetic field, field gradients andradiofrequency energy to excite protons and thereby make an image of themobile protons in water and fat. MRI has found many applications inimaging the central nervous system, but abdominal applications havelagged seriously behind. One reason that abdominal MRI has not beenutilized more extensively has been the absence of a suitable MRIcontrast agent for the gastrointestinal tract. Computed tomography (CT)is used more commonly for abdominal imaging in part because suitablecontrast agents, chiefly barium and iodine compounds, are available foruse in such imaging.

MRI contrast agents primarily act by affecting T1 or T2 relaxation ofwater protons. Contrast agents generally shorten T1 and/or T2. Whencontrast agents shorten T1, this increases signal intensity on T1weighted images. When contrast agents shorten T2, this decreases signalintensity particularly on T2 weighted pulse sequences. To date severalprototype gastrointestinal MRI contrast agents have been developed toassist abdominal MRI, but none of these have been altogethersatisfactory.

For example, iron oxides which are strong T2 relaxation agents have beenused as negative gastrointestinal MRI contrast agents to decrease signalintensity in the gastrointestinal tract. These agents, whichpredominantly affect T2, have the disadvantages of magneticsusceptibility artifacts which occurs as a result of the drastic effectson local magnetic homogeneity (magnetic susceptibility) caused by theseagents. Magnetic susceptibility artifacts make it difficult to assessthe bowel wall, bowel mesentery and adjacent structures.

The paramagnetic MRI contrast agent gadolinium-DTPA has also been testedas a positive gastrointestinal MRI contrast agent to increase signalintensity on T1 weighted images, but this agent has the drawback thatdecomplexation and release of free gadolinium ion may occur in thegastrointestinal tract which can be quite toxic. Furthermore,gadolinium-DTPA is relatively expensive.

Ferric iron has also been experimented with as an oral gastrointestinalMRI contrast agent. Ferric iron has been administered in the form offerric ammonium citrate wherein the paramagnetic Fe⁺³ iron relaxes thewater in the bowel to make the bowel bright on T1 weighted images.Ferric ammonium citrate is quite inexpensive but the resultantgastrointestinal MRI contrast agent has been suboptimally useful. Toobtain reasonable contrast enhancement, a relatively high dose of ferriciron is required, and some of this iron is absorbed as it passes downthe gastrointestinal tract. Absorption of the iron creates two problems.First, absorption of the iron may cause problems with iron toxicity andiron overload. Second, as the iron is absorbed from the gastrointestinaltract, the concentration of the contrast agent decreases and the degreeof contrast enhancement is much less in the distal bowel.

The ideal contrast agent for the gastrointestinal tract would affectboth T1 and T2, causing the tract lumen to appear bright on T1 weightedimages and dark on T2 weighted images. Tumors and other pathologictissues generally have a long T1 and a long T2, which is to say thatthese pathologic tissues appear dark on T1 weighted images and bright onT2 weighted images. If the lumen could be filled with contrast materialwhich appeared bright on T1 weighted images and dark on T2 weightedimages, it would then be easy to differentiate a normal gastrointestinaltract from any adjacent abnormal tissues. The ideal contrast agent wouldalso serve to minimize any toxicity problems, and be relativelyinexpensive.

The need is great for new gastrointestinal MRI contrast agents, havingsome or all of the aforementioned qualities. The present invention isdirected to achieving this important end.

SUMMARY OF THE INVENTION

The present invention is directed to magnetic resonance imaging, andmore particularly to the use of a contrast medium comprising acombination of at least one polyphosphorylated aliphatic or alicycliccompound of five or more carbon atoms and at least one paramagnetic ionto image the gastrointestinal region of a patient.

Specifically, the present invention pertains to methods of providing animage of the gastrointestinal region of a patient comprising (i)administering to the patient the aforementioned contrast medium, and(ii) scanning the patient using magnetic resonance imaging to obtainvisible images of that region.

The present invention is further directed to methods for diagnosing thepresence of diseased tissue in the gastrointestinal region of a patientcomprising (i) administering to the patient the aforementioned contrastmedium, and (ii) scanning the patient using magnetic resonance imagingto obtain visible images of any diseased tissue in the patient.

The present invention also provides diagnostic kits for gastrointestinalimaging which include the subject contrast medium.

The polyphosphorylated compounds, when employed in combination withparamagnetic ions, provide highly effective and relatively inexpensivecontrast enhancement agents for gastrointestinal magnetic resonanceimaging. Unlike many of the gastrointestinal agents of the prior art,where contrast enhancement has been either positive or negative, thepresent invention provides a contrast agent that may impart bothpositive and negative contrast. The polyphosphorylated compounds used inthe present invention serve to effectively bind the paramagnetic ion,thereby minimizing the potential for absorption of potentially toxicparamagnetic ion throughout the gastrointestinal region. This means thatnot only can a lower dose of paramagnetic ion be used for contrastenhanced magnetic resonance imaging than would be possible withoutpolyphosphorylated compounds, but also that a relatively uniformconcentration of the ion can be achieved throughout the gastrointestinaltract. Moreover, the polyphosphorylated compounds utilized in thepresent invention have been found to enhance the relaxivity of theparamagnetic ions. In the case of the combination of thepolyphosphorylated compound inositol hexaphosphate with the paramagneticagent ferric iron, for example, relaxivity in the gastrointestinalregion was found to increase by a factor of almost three-fold incomparison with the use of ferric iron alone. Simultaneously, theabsorption of ferric iron in the gastrointestinal tract was found todecrease by over 80%. Heretofore, the use of such agents forgastrointestinal imaging and the safe and highly effective contrastenhancement achieved thereby, was neither disclosed nor suggested.

These and other aspects of the invention will become more apparent fromthe following detailed description when taken in conjunction with thefollowing figures.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1. The T1 values for solutions of ferric citrate in combinationwith a concentration of 0.33 or 1.0 millimolar inositol hexaphosphate,and the T1 values for solutions of ferric citrate without inositolhexaphosphate, are shown. In each solution, 2 weight % cellulose waspresent, as well as 0.25 weight % xanthum gum. As the figure reveals,there is an approximately three-fold effect on the T1 relaxation timescaused by the ferric iron and inositol hexaphosphate combination, ascompared to ferric iron without the inositol hexaphosphate.

FIG. 2. The T1 relaxation time of water alone, and water with 0.33millimolar inositol hexaphosphate are shown. The addition of inositolhexaphosphate to the water has no appreciable effect on T1 relaxivity,at most causing a mild prolongation of T1, but showing no improvement inT1 relaxivity.

FIG. 3a and 3b. The T1 (FIG. 3a) and T2 (FIG. 3b) relaxation times of1.0 millimolar concentration of ferric iron in combination with inositolhexaphosphate concentrations varying between 0.1 and 2.0 millimolar areshown. The optimum ratio of ferric iron to inositol hexaphosphateappears to be about 1 to 1 for maximizing relaxivity.

FIGS. 4a and 4b. The T1 (FIG. 4a) and T2 (FIG. 4b) relaxation times of 1millimolar solutions of ferric iron with 1 millimolar inositolhexaphosphate at varying pHs are shown. The T1 and T2 relaxation timesare appreciably shorter between pH 4 and pH 10, which reflects astronger interaction with inositol hexaphosphate with ferric iron overthis range of pH values.

FIG. 5. The T2 signal intensity on magnetic resonance images is shownfor phosphate buffered saline (PBS) and a solution of 1 millimolarferric iron with 1 millimolar inositol hexaphosphate at a constant TRand at echo times varying from 50 to 250 milliseconds. As shown in thisfigure, the signal intensity of the ferric iron/inositol hexaphosphatecontrast agent falls much more rapidly than saline. On a T2 weightedimage of the gastrointestinal tract containing concentrations of ferriciron/inositol hexaphosphates greater than 1 millimolar (not shown),there was an appreciable decrease in signal intensity of the fluid inthe bowel.

FIGS. 6a and 6b. The effect on T1 of ferric iron with inositolhexaphosphate, and ferric iron with inositol hexaphosphate and 2 weight% of cellulose (FIG. 6a), and the effect on T2 of ferric iron with 2weight % of cellulose and 0.25 weight % xanthan gum, and ferric ironwith inositol hexaphosphate and 2 weight % cellulose and a 0.25 weight %xanthan gum (FIG. 6b) is shown. The addition of 2 weight % cellulose toa solution of ferric iron and inositol hexaphosphate causes a slightincrease in T1 relativity. The presence of cellulose has an even greatereffect on T2 relativity.

FIG. 7. The stacking phenomenon that is believed to occur where aninositol hexaphosphate is combined with a paramagnetic ion is shown. Inaccordance with this theory, the inositol hexaphosphate is believed tobind to a paramagnetic ion, which in turn binds to anotherpolyphosphorylated compound, which in turn may bind to anotherparamagnetic ion, which in turn may bind to another phosphorylatedcompound, and so on, forming a copolymer of varying length of thephosphorylated compound and the paramagnetic ion. The stacking of threeinositol hexaphosphate compounds with the paramagnetic ion iron is shownfor illustration purposes.

DETAILED DESCRIPTION OF THE INVENTION

Any of the wide variety of biocompatible polyphosphorylated aliphatic oralicyclic compounds of at least five carbon atoms that are known in theart may be employed in the methods and kits of the present invention.The term biocompatible, used throughout the specification, is employedin its conventional sense, that is, to denote compounds that do notsubstantially interact with the tissues, fluids and other components ofthe body in an adverse fashion in the particular application ofinterest. The term polyphosphorylated, used in connection with thecompounds of the present invention, denotes compounds containing two ormore phosphate (e.g., PO₄ ⁻³, PO₃ H⁻¹ or PO₂ H⁻²) substituents. Theterms aliphatic and alicyclic, used herein, are employed in theconventional sense, that is, aliphatic denotes herein organic compoundshaving an open chain structure, and alicylic denotes herein organiccompounds having a saturated ring structure.

Preferably, the polyphosphorylated compounds of the invention contain atleast three phosphate groups, more preferably at least four phosphategroups, even more preferably at least five phosphate groups, and mostpreferably at least six phosphate groups. In the case ofpolyphosphorylated alicyclic compounds, it is preferable that thephosphate groups be located both above and below the plane of the ring,that is, one pair of phosphate groups are located trans to one another.Also, in the case of aliphatic compounds, the compound is preferably atleast about ten carbon atoms in length, more preferably at least aboutfifteen carbon atoms in length, even more preferably at least abouttwenty carbon atoms in length, and most preferably at least about thirtycarbon atoms in length. Preferably, the molecular weight of thephosphorylated aliphatic compound would be in the range of about 20,000to about 100,000, most preferably between about 50,000 and about100,000. In the case of alicyclic compounds, the cyclic compoundpreferably contains at least six carbon atoms in its ring structure, andthere may, if desired, be more than one carbon ring structure in tandem,that is dimers (e.g., disaccharides), trimers, oligomers, and polymers(e.g., polysaccharides). As will be readily apparent to those skilled inthe art, once armed with the present disclosure, there are a number ofpolyphosphorylated compounds that can be effectively employed in thepresent invention, and any of such compounds, as well as anycombinations thereof are intended to be within the scope of the presentinvention.

Suitable biocompatible polyphosphorylated aliphatic compounds of atleast five carbon atoms include biocompatible aliphatic compounds whichmay be polyphosphorylated, such as biocompatible aliphatic compoundscontaining hydroxyl, keto, amino, or unsaturated groups. Suchbiocompatible aliphatic compounds include, but are not limited to,polyphosphorylated polyvinyl alcohol, polyphosphorylated polyethyleneglycol, polyphosphorylated polypropylene glycol, polyphosphorylatedpolystyrene, polyphosphorylated polyacrylic acid, polyphosphorylatedpolymethacrylic acid, polyphosphorylated copolymers of acrylic andmethacrylic acid, and various polyphosphorylated polyolefins.

Suitable biocompatible polyphosphorylated alicyclic compounds of atleast five carbon atoms include, but are not limited to:polyphosphorylated sugar alcohols such as polyphosphorylated inositol,polyphosphorylated mannitol, polyphosphorylated sorbitol,polyphosphorylated pentaerythritol, polyphosphorylated galacitol,polyphosphorylated adonitol, polyphosphorylated arabitol, andpolyphosphorylated xylitol; polyphosphorylated monosacharides such aspolyphosphorylated glucose, polyphosphorylated fructose,polyphosphorylated mannose, polyphosphorylated idose, polyphosphorylatedgalactose, polyphosphorylated allose, polyphosphorylated altrose, andpolyphosphorylated arabinose; polyphosphorylated disaccharides such aspolyphosphorylated sucrose, polyphosphorylated maltose,polyphosphorylated cellobiose, and polyphosphorylated lactose; andpolyphosphorylated polysaccharides such as polyphosphorylated cellulose,polyphosphorylated agarose, polyphosphorylated lignan, andpolyphosphorylated chitin.

For reasons of diagnostic efficacy and biocompatibility,polyphosphorylated inositol and/or polyphosphorylated cellulosecompounds are most preferred.

Such polyphosphorylated aliphatic and alicyclic compounds can be easilyprepared from readily available starting materials, using conventionalsynthesis techniques, as will be apparent to those skilled in the art.Conventional phosphorylating agents include such compounds asphosphorous chloride (mono-, di-, or tri-), phosphorous bromide (mono-,di-, or tri-), phosphorous oxychloride (mono-, di-, or tri-),phosphorous oxybromide (mono-, di-, or tri-), phosphorous pentaoxide,phosphoric acid, phosphorous acid, and the anhydrides thereof,pyrophosphoric acid, aziridine phosphine oxide, choloroalkylphosphonicacid or its derivatives, and phosphorous acid ester.

In accordance with one preparatory protocol, starting with an aliphaticor alicyclic compound containing one or more hydroxyl groups, forexample, phosphorylation can be easily carried out by suspending thestarting material in, for example, chloroform, then adding a phosphoricester monochloride compound to the suspension, preferably dropwise.Suitable phosphoric ester monochloride compounds include ClP(O) (OR)₂,wherein R is selected from, inter alia, C(O)CH₃, C(O)H, CH₃, C₂ H₅, C₃H₇, C₄ H₉, and CH₂ C₆ H₅. The resulting phosphorylated compound can thenbe treated with water to hydrolyze to the corresponding phosphonic acidderivatives. Such hydrolyzed derivatives are included within the scopeof the phrase phosphorylated compounds herein.

In another method for preparing the phosphorylated aliphatic andalicylic compounds of the invention, the urea catalyzed phosphoric acid(or phosphorous acid) phosphorylation procedures may be convenientlyutilized. In this procedure, aliphatic or alicyclic compounds are soakedwith mixtures of urea and phosphoric acid (or phosphorous acid), thenheated to 120° C., or higher.

In a further method, phosphorylation of compounds such as polyvinylalcohol can be carried out by dissolving the polyvinyl alcohol in anorganic solvent such as pyridine, dimethylformamide, ordimethylsulfoxide with triethylamine, and then addingdialkyloxyphosphoric monochloride ((RO)₂ POCl) to the solution. Afterphosphorylation, the solutions are hydrolyzed by adding water andacidifying with hydrochloric acid. Unbound phosphorous is then removedby passing the solution through a column filled anion exchanger.

Other methods of phosphorylating various compounds to producephosphorylated compounds within the scope of the present invention aredisclosed, for example, in Sander et al., J. Macromol. Sci., Rev.Macromol. Chem., Vol. 2, pp. 57-72 (1968), Leonard et al., J. App.Polymer Sci., Vol. V, pp. 157-162 (1961), Leonard et al., J. of PolymerSci., Vol. 55, pp. 799-810 (1961), Schroeder et al., J. of Polymer Sci.,Vol. XLVII, pp. 417-433 (1960), Kennedy et al., J. Appl. Chem., Vol. 8,pp. 459-464 (1958), Marvel et al., J. of Polymer Sci., Vol. VIII, pp.495-502 (1952), the disclosures of each of which are hereby incorporatedherein by reference, in their entirety.

As a general matter, it is believed that the higher the degree ofphosphorylation in the aliphatic and alicyclic compounds, the morediagnostically effective and less toxic is the contrast medium whenemployed as a gastrointestinal contrast agent. For example, although allpolyphosphorylated inositol compounds can be employed in the imagingmethods of the invention, inositol hexaphosphate is preferred overinositol pentaphosphate, which in turn is preferred over inositoltetraphosphate, which in turn is preferred over inositol triphosphate.It is also believed that, for reasons of diagnostic efficacy, a largercarbon chain is preferred over a smaller one. Thus, for example, a sixcarbon containing alicyclic compound such as inositol polyphosphate ispreferred over a five carbon containing alicyclic compound such asarabinose polyphosphate, and a thirty carbon aliphatic compound such asa thirty carbon polyvinyl alcohol is preferred over a twenty carbonaliphatic compound such as a twenty carbon polyvinyl alcohol.

As noted above, the polyphosphorylated aliphatic and alicyclic compoundsare employed in combination with paramagnetic ions. Suitableparamagnetic ions include, but are not limited to, compounds comprisingtransition, lanthanide and actinide elements, and any of the suitableparamagnetic ions, and any combinations thereof, are intended to bewithin the scope of the present invention. Preferable of such elementsare Gd(III), Mn(II), Cu(II), Cr(III), Fe(II), Fe(III), Co(II), Er(II),Ni(II), Eu(III), Yb(III) and Dy(III). More preferably, the elements areFe(III), Gd(III), Mn(II), Cu(II), Cr(III), Yb(III) and Dy(III),especially Fe(III) and Mn(II). The paramagnetic ions may be added, ifdesired, as a salt, such as, for example, in the case of ferric iron(Fe(III)), as ferric citrate, ferric chloride, ferric acetate, ferricglycerophosphate, ferric sulfate, ferric phosphate, and ferric ammoniumphosphate. The preferable paramagnetic ions are Fe(III), added in theform of ferric citrate, and Mn(II), added in the form of manganesechloride, manganese acetate, manganese sulfate or manganese phosphate.

As noted above, combinations of paramagnetic ions are within the scopeof the present invention. Choice of appropriate combinations ofparamagnetic ions can increase the ultimate relaxivity and contrastenhancement of the contrast media of the present invention. By way ofexample, an extremely good combination of paramagnetic ions for thecontrast media of the present invention are manganese and iron. Thiscombination takes into account the fact that manganese is a bettercontrast agent than iron, that both are absorbed from thegastrointestinal tract through similar receptors, and that iron ispreferentially absorbed over manganese, a competitive absorption type ofsituation. The inclusion of the iron would thus serve to minimize anyabsorption of the more active manganese, resulting in a better contrastmedium. This phenomenon is shown in Examples 6 and 7, below.

As one skilled in the art would recognize, wide variations in theamounts of the polyphosphorylated compound and paramagnetic ion can beemployed in the methods and kits of the invention, with the preciseamounts varying depending upon such factors as the mode ofadministration (e.g., oral, rectal), and the specific portion of thegastrointestinal tract for which an image is sought (e.g., theesophagus, stomach, rectum, etc). Preferably, however, the paramagneticions are present in a concentration between about 1 to about 4millimolar. The polyphosphorylated compounds are preferably present in amolar concentration of between about 0.1 and about 3 times the molarconcentration of the paramagnetic ions. Most preferably, the molarconcentration of the polyphosphorylated alicyclic compounds is about 1.0times the molar concentration of the paramagnetic ions. Most preferably,the molar concentration of the phosphorylated aliphatic compounds isbetween about 0.1 and about 0.5 times the molar concentration of theparamagnetic ions. The volume of contrast agent administered to thegastrointestinal tract is preferably between about 500 to about 1000 cc,and is supplied in the form of an aqueous solution.

The polyphosphorylated compounds and paramagnetic ion compositions maybe employed alone, if desired, as a contrast medium for gastrointestinalmagnetic resonance imaging. Alternatively, if desired, they may beemployed in conjunction with other biocompatible synthetic or naturalpolymers. By the phrase in conjunction with it is meant that thepolymers may be simply added to the polyphosphorylated compound andparamagnetic ion mixture, or alternatively may be bound to thepolyphosphorylated compound by a covalent linkage, the binding beingaccomplished using conventional methodology, such as, for example, bythe procedures described in Breitenbach et al., in Phytic Acid:Chemistry and Applications, pp. 127-130, Graf, ed., (Pilatus Press,Minneapolis, Minn. 1986), the disclosures of which are incorporatedherein by reference in their entirety. Exemplary suitable syntheticpolymers include polyethylenes (such as, for example, polyethyleneglycol), polyoxyethylenes (such as, for example, polyoxyethyleneglycol), polypropylenes (such as, for example, polypropylene glycol),pluronic acids and alcohols, polyvinyls (such as, for example, polyvinylalcohol), and polyvinylpyrrolidone. Exemplary suitable natural polymersinclude polysaccharides. Such polysaccharides include, for example,arabinans, fructans, fucans, galactans, galacturonans, glucans, mannans,xylans (such as, for example, inulin), levan, fucoidan, carrageenan,galactocarolose, pectic acid, amylose, pullulan, glycogen, amylopectin,cellulose, carboxylmethylcellulose, hydroxypropyl methylcellulose,dextran, pustulan, chitin, algin, agarose, keratan, chondroitin,dermatan, hyaluronic acid and alginic acid, and various otherhomopolymers or heteropolymers such as those containing one or more ofthe following aldoses, ketoses, acids or amines: erythrose, threose,ribose, arabinose, xylose, lyxose, allose, altrose, glucose, mannose,gulose, idose, galactose, talose, erythrulose, ribulose, xylulose,psicose, fructose, sorbose, tagatose, glucuronic acid, gluconic acid,glucaric acid, galacturonic acid, mannuronic acid, glucosamine,galactosamine and neuraminic acid. Algin has been found to be aparticularly useful polymer to use in conjunction with the contrastmedia of the invention because this compound, which binds about 300times its weight in water and has a slippery texture, hastensgastrointestinal transit and promotes gastric emptying. Its use inconjunction with the contrast media of the invention is illustrated inExample 7, below. As those skilled in the art will recognize armed withthe present disclosure, such polymers may be added in varying amounts,as desired.

The polyphosphorylated compounds and paramagnetic ion compositions ofthe invention may also be employed, if desired, with other agents whicheffect T2 relaxation such as bismuth, barium, kaolin, atapulgite, ferricoxide (either uncoated or coated with, for example, dextran, algin,cellulose, etc., for improving the suspension of the ferric oxideparticles), or stabilized gas (stabilized with, for example,microspheres of polyvinylidene acrylonitrile copolymers or acrylonitrilepolymers of about 20 to about 100 microns in size).

If desired, in addition, the contrast medium of the present inventionmay be utilized with biocompatible anti-oxidant compounds. Suitableanti-oxidants include vitamin C (ascorbic acid), vitamin E (tocopherol),and retinoic acid. Other suitable anti-oxidants will be readily apparentto those skilled in the art. The anti-oxidants may be employed incertain applications to keep paramagnetic ions, such as manganese, intheir more paramagnetically effective reduced state, that is, forexample, in the Mn(II) state, rather than the Mn(III) state.

The contrast medium utilized in the gastrointestinal applications of thepresent invention may also be employed with conventional biocompatibleanti-gas agents. As used herein the term anti-gas agent is a compoundthat serves to minimize or decrease gas formation, dispersion and/oradsorption. A number of such agents are available, including antacids,antiflatulents, antifoaming agents, and surfactants. Such antacids andantiflatulents include, for example, activated charcoal, aluminumcarbonate, aluminum hydroxide, aluminum phosphate, calcium carbonate,dihydroxyaluminum sodium carbonate, magaldrate magnesium oxide,magnesium trisilicate, simethicone, sodium carbonate, loperamidehydrochloride, diphenoxylate, hydrochloride with atropine sulfate,Kaopectate™ (kaolin) and bismuth salts. Suitable antifoaming agentsuseful as anti-gas agents include simethicone, protected simethicone,siloxyalkylene polymers, siloxane glycol polymers,polyoxypropylene-polyoxyethylene copolymers, polyoxyalkylene amines andimines, branched polyamines, mixed oxyalkylated alcohols, finely dividedsilica either alone or mixed with dimethyl polysiloxane, sucroglycamides(celynols), polyoxylalkylated natural oils, halogenatedsilicon-containing cyclic acetals, lauryl sulfates, 2-lactylic acidesters of unicarboxylic acids, triglyceride oils. Particles of polyvinylchloride or silica may also function as anti-foaming agents in thesubject invention. Suitable surfactants include perfluorocarbonsurfactants, such as, for example, DuPont Zonyl™ perfluoroalkylsurfactants known as Zonyl™ RP or Zonyl™ NF, available from DuPont,Chemicals and Pigments Division, Jackson Laboratory, Deepwater, N.J.08023. Of course, as those skilled in the art will recognize, anyanti-gas agents employed must be suitable for use within the particularbiological system of the patient in which it is to be used. Theconcentration of such anti-gas agents may vary widely, as desired, aswill be readily apparent to those skilled in the art. Typically,however, such agents are employed in concentrations of between about 20and about 2000 ppm, most preferably in concentrations between about 50and about 1000 ppm.

The present invention is useful in imaging the gastrointestinal regionof a patient and in diagnosing the presence of diseased tissue in thatregion. The imaging process of the present invention may be carried outby (a) administering to a patient a contrast medium comprising (i) atleast one polyphosphorylated compound comprising at least five carbonatoms, and (ii) at least one paramagnetic ion, and (b) then scanning thepatient using magnetic resonance imaging to obtain visible images of thegastrointestinal region of a patient and/or of any diseased tissue inthat region. The phrase gastrointestinal region or gastrointestinaltract, as used herein, includes the region of a patient defined by theesophagus, stomach, small and large intestines, and rectum. The patientcan be any type of mammal, but most preferably is a human.

As one skilled in the art would recognize, administration may be carriedout in various fashions, such as orally, rectally, intravascularly,using a variety of dosage forms. Since the region to be scanned is thegastrointestinal region, administration of the contrast medium of theinvention is preferably carried out orally or rectally. The usefuldosage to be administered and the particular mode of administration willvary depending upon the age, weight and the particular mammal to bescanned, the particular portion of the gastrointestinal region to bescanned, and the particular contrast medium to be employed. Typically,dosage is initiated at lower levels and increased until the desiredcontrast enhancement is achieved. If desired, a multiple dosing methodof delivering the contrast agents of the invention may be employed toprovide uniformity of enhancement throughout the gastrointestinal tract,such as is illustrated in Examples 6 and 7, below.

Kits useful for magnetic resonance imaging of the gastrointestinalregion in accordance with the present invention comprise at least onephosphorylated compound comprising at least five carbon atoms and atleast one paramagnetic ion, in addition to conventional proton magneticresonance imaging kit components. Such conventional proton magneticresonance imaging kit components include those described above, as wellas other components which will be readily apparent to those skilled inthe art, once armed with the present disclosure, such as those describedin Weinmann et al., U.S. Pat. No. 4,719,098, the disclosures of whichare hereby incorporated by reference in their entirety. Exemplarycomponents which may be employed in the kit in addition to thephosphorylated compounds comprising at least five carbon atoms and theparamagnetic ions, include polymers, anti-oxidants, anti-gas agents,various T2 relaxation agents, osmolality raising agents, viscosity andbulking agents, buffering agents, and gastrointestinal transit agents todecrease gastrointestinal transit time and increase rate ofgastrointestinal emptying.

Such kit componenets as polymers, anti-oxidants, anti-gas agents, and T2relaxation agents have been described in detail above.

Suitable osmolality raising agents include polyols and sugars, forexample, mannitol, sorbitol, arabitol, xylitol, glucose, sucrose,fructose, and saccharine, with mannitol and sorbitol being mostpreferred. The concentration of such osmolality raising agents may vary,as desired, however, generally a range of about 5 to about 70 g/l,preferably about 30 to about 50 g/l of the contrast medium. Suchcompounds may also serve as sweeteners for the ultimate formulation, ifdesired.

Suitable viscosity and bulking agents include the polymers described indetail above, as well as other agents which are well known in the art toprovide viscosity and bulking. Particularly useful are alginates,xanthan gum, guar, pectin, tragacanth, bassorin, karaya, gum arabic,casein, gelatin, sodium carboxymethylcellulose, methylcellulose,methylhydroxycellulose, bentonite, collodial silicic acid, and variousanti-diarrhetic preparations. Such compounds may be employed in varyingamounts, as those skilled in the art would recognize, but preferably areemployed in amounts of about 2 to about 40 g/l, preferably about 10 toabout 30 g/l of the contrast medium.

Buffering agents, that is buffers, buffer mixtures and bases, may beutilized to stabilize the phosphorylated compound and paramagnetic ioncomplex with respect to the acidic stomach content. Such bufferingagents include tris(hydroxymethyl)aminomethane(2-amino-2-hydroxymethyl-1,3-propanediol, trometamol), sodium dihydrogenphosphate/disodium hydrogen phosphate, citric acid/disodium phosphate,etc., with trometamol being preferred. Other suitable buffering agentsinclude all physiologically compatible organic and inorganic bases,e.g., sodium carbonate, calcium carbonate, amino sugars (e.g.,glucosamine), amino alcohols (e.g., methylglucamine), amino acids (e.g.,arginine, lysine), and the like. The desired pH range is about 3 toabout 9 pH units, preferably about 4 to about 9 pH units in thegastrointestinal region to be imaged. In view of the strong bindingcapabilities, relatively speaking, of the phosphorylated compounds, suchbuffering agents may be unnecessary. If employed, the buffering agentsmay be used in varying amounts, as will be readily apparent to thoseskilled in the art, generally in concentrations between about 5 andabout 40 mmol/l.

Gastrointestinal transit agents to decrease gastrointestinal transittime and increase rate of gastrointestinal emptying include algin, aswell as many of the compounds listed above as viscosity and bulkingagents, with algin being most preferred. The amount of such agents will,of course, vary as those skilled in the art will recognize, butgenerally will be employed in an amount of between about 5 and about 40mmol/l.

The magnetic resonance imaging techniques which are employed areconventional and are described, for example, in D. M. Kean and M. A.Smith, Magnetic Resonance Imaging: Principles and Applications,(Williams and Wilkins, Baltimore 1986). Contemplated MRI techniquesinclude, but are not limited to, nuclear magnetic resonance (NMR) andelectronic spin resonance (ESR). The preferred imaging modality is NMR.

Phosphorylated compounds utilized in the gastrointestinal imagingmethods of the present invention have been found to enhance therelaxivity of the paramagnetic ions with which they are combined. In thecase of the combination of the phosphorylated alicyclic compoundinositol hexaphosphate with the paramagnetic agent ferric iron, forexample, relaxivity in the gastrointestinal region was found to increaseby a factor of almost three-fold in comparison with the use of ferriciron alone (FIG. 1). As shown in FIG. 6, the T2 relaxation effects maybe further enhanced by using a combination of inositol hexaphosphate andcellulose with the paramagnetic ion ferric iron. The combination ofphosphorylated compound and paramagnetic ion of the present inventionmay act to both increase signal intensity on T1 weightedgastrointestinal images and decrease signal intensity on T2 weightedgastrointestinal images, providing both positive and negative contrast,an ideal characteristic for a gastrointestinal contrast agent.

The greatly enhanced relaxivity resulting from, for example, thecombination of ferric iron with inositol hexaphosphate ingastrointestinal imaging, has never been reported and is indeedunexpected. Generally, when a paramagnetic ion is bound by acomplexation agent (a ligand), there is a reduction in relaxivity causedby shielding of the paramagnetic centers from water. An example isgadolinium-DTPA wherein the complex has about one-half the relaxivity offree gadolinium ion. It is unexpected that a phosphorylated compound,such as, for example, inositol hexaphosphate, might improve therelaxivity of, for example, iron or manganese so dramatically.

Although not intending to be bound by any theory or mechanism ofoperation, it is believed that the phosphorylated compounds employed inthe gastrointestinal imaging methods of the invention may act to improvethe relaxivity of the paramagnetic ions by slowing the correlation timeof the iron, that is, by slowing down the rate at which the iron istumbling in solution, rather than by some cross relaxation mechanism. Asshown in FIG. 2, inositol hexaphosphate appears to have little, if any,significant effect on the relaxation rate of water. If cross-relaxationwere the dominant mechanism responsible for the increase in relaxivitywith the iron plus inositol hexaphosphate contrast medium, then theinositol hexaphosphate alone should have some measure of relaxationeffect on water. Moreover, when the inositol hexaphosphate is added tothe solution of ferric iron as shown in FIG. 1, there is a dose relatedeffect on relaxivity. This change in relaxivity may reflect the inositolhexaphosphate binding the iron and slowing the tumbling rates, resultingin a more favorable correlation time of the iron such that therelaxivity of the iron is enhanced. Further to this theory, it isbelieved that when the paramagnetic ions are added to thepolyphosphorylated aliphatic and alicyclic compounds, a stackingphenomenon tends to occur wherein a polyphosphorylated compound binds toa paramagnetic ion, which in turn binds to another polyphosphorylatedcompound, which in turn binds to another paramagnetic ion, which in turnbinds to another phosphorylated compound, and so on, forming in essencea "copolymer" of the phosphorylated compound "monomer" and theparamagnetic ion "monomer". This stacking phenomenon ("copolymer"formation) is schematically illustrated in FIG. 7. It is believed thatthe stacking phenomenon may, at least in part, account for highlydiagnostically effective properties of the preferred compounds of thepresent invention, providing slower tumbling rates, more favorablecorrelation times and enhanced relaxivity.

Another advantage of the present invention is that a smaller amount ofthe paramagnetic ion is absorbed in the gastrointestinal tract when thephosphorylated compounds are employed in combination therewith. Forexample, the absorption of ferric iron in the gastrointestinal tract wasfound to decrease by over 80% when employed in combination with inositolhexaphosphate. One benefit to patients utilizing the present inventionis that a lower dose of contrast agent is needed to produce clinicallyuseful contrast enhancement of the gastrointestinal tract. It ispossible to achieve the same pattern of enhancement in thegastrointestinal region using paramagnetic agent without aphosphorylated compound, but an approximately three-fold higherconcentration of paramagnetic ion may have to be utilized, and such ahigh amount of paramagnetic ion may prove toxic. Moreover, where only aparamagnetic agent is used, absorption of the paramagnetic ion may be aproblem, not only in causing toxicity, but may also result in diminishedcontrast in the distal portion of the gastrointestinal tract.

The present invention is further described in the following Examples.These Examples are not to be construed as limiting the scope of theappended claims.

EXAMPLES Example 1

A contrast medium was formulated by preparing a 900 cc aqueous solutioncontaining 120 mg of ferric ammonium citrate with 720 mg of inositolhexaphosphate (1 to 1 molar ratio of ferric iron to inositolhexaphosphate), and was orally ingested by a human test volunteer. T1and T2 weighted spin echo images of the abdomen were obtained usingmagnetic resonance imaging. High signal intensity was observed on boththe T1 and T2 weighted images.

Example 2

A contrast medium was formulated by preparing a 900 cc aqueous solutioncontaining 200 mg of ferric ammonium citrate with 1200 mg of inositolhexaphosphate (1 to 1 molar ratio of ferric iron to inositolhexaphosphate), and was orally ingested by a human test volunteer.Images were taken of the gastrointestinal region of the volunteer usingmagnetic resonance imaging, both pre and post ingestion. The imagesrevealed greatly enhanced contrast of the gastrointestinal region postingestion, particularly in the stomach and small intestine (includingthe duodenum) areas, with high signal intensity on the T1 weightedimages (high positive contrast) and somewhat low signal intensity(slight negative contrast) on the T2 weighted images in thegastrointestinal lumen being observed.

Example 3

The procedures of Example 2 were substantially repeated, except that inaddition to the ferric ammonium citrate and inositol hexaphosphate, 9 gof phosphorylated cellulose (1% by weight) was added to the solution.The results, following imaging, were substantially similar to Example 1,except that even lower signal intensity (higher negative contrast) onthe T2 weighted images was observed.

Example 4

A contrast medium was formulated by preparing a 1 liter aqueous solutioncontaining 5.6 mg of manganese (II) chloride (approximately 0.1 mMmanganese) and 0.3 mM inositol hexaphosphate (an approximate 1 to 3molar ratio of manganese to inositol hexaphosphate), and was orallyingested by a human test volunteer. Images were taken of thegastrointestinal region of the volunteer using magnetic resonanceimaging. The resultant images showed slight high signal intensity on theT1 weighted images, but no appreciable contrast on the T2 weightedimages.

Example 5

Three separate contrast media were formulated by preparing aqueoussolutions containing, respectively, 20, 30 and 40 mg of manganese (II)per liter, each containing a three-fold excess of inositol hexaphosphatein an iso-osmotic solution. The osmolality of the solution was adjustedto about 300 mosm using sorbitol and polyethylene glycol. In addition,approximately 2 grams of vitamin C per liter was added to each solutionto minimize oxidation of the Mn (II) to the less favorable relaxationstate of Mn (III). Xanthan gum (0.25 weight %) was also added. Eachsolution was then ingested by a different human test volunteer, andimages were taken of the gastrointestinal region using magneticresonance imaging. In each case, the solution showed high signalintensity on the T1 weighted images of the bowel with the region ofenhancement extending from the stomach into the distal small bowel. Onthe T2 weighted images, the preparations with 30 and 40 mg of manganeseper liter showed low signal intensity on the T2 weighted images whichwas more pronounced for the concentration of 40 mg per liter ofmanganese. Even at a concentration of 40 mg manganese per liter,however, the distal portion of the small bowel still appeared bright onthe T2 weighted images.

Example 6

Three separate contrast media were formulated by preparing aqueoussolutions in three separate bottles of 300 cc each. Bottle #1 contained60 mg of manganese (II) and 60 mg of iron (II) per liter, and athree-fold excess inositol hexaphosphate in an iso-osmotic solution.Bottle #2 contained 50 mg of manganese (II) and 50 mg of iron (II) perliter, and a three-fold excess inositol hexaphosphate in an iso-osmoticsolution. Bottle #3 contained 40 mg of manganese (II) and 40 mg of iron(II) per liter, and a three-fold excess inositol hexaphosphate in aniso-osmotic solution. The contents of each of the three bottles werethen ingested over a short period of time by a human test volunteer, inthe order of bottle #1 first, then bottle #2, and finally bottle #3, andimages were taken of the gastrointestinal region using magneticresonance imaging. Enhancement was found to be much more uniform withpositive contrast on T1 and negative contrast on T2 weighted images.Enhancement in the colon area, however, was less than desired.

Example 7

Three separate contrast media were formulated by preparing aqueoussolutions in three separate bottles of 300 cc each. Bottle #1 contained75 mg of manganese (II), 75 mg of iron (II) per liter, a three-foldexcess inositol hexaphosphate, 2 g vitamin C, 0.75 weight % algin, and0.25 weight % xanthan gum. Bottle #2 contained 60 mg of manganese (II),60 mg of iron (II) per liter, a three-fold excess inositolhexaphosphate, 2 g vitamin C, 0.5 weight % algin, and 0.25 weight %xanthan gum. Bottle #3 contained 50 mg of manganese (II), 50 mg of iron(II) per liter, a three-fold excess inositol hexaphosphate, 2 g vitaminC, 0.3 weight % algin, and 0.25 weight % xanthan gum. Osmolality of eachsolution was adjusted to about 300 mosm using 1% sorbitol and 2%polyethylene glycol in each bottle. The contents of each of the threebottles were then ingested over a short period of time by a human testvolunteer, in the order of bottle #1 first, then bottle #2, and finallybottle #3, and images were taken of the gastrointestinal region usingmagnetic resonance imaging. Enhancement was found to be even much moreuniform than in Example 6, with positive contrast on T1 and negativecontrast on T2 weighted images. Enhancement was visualized throughoutthe entire gastrointestinal tract.

Various modifications of the invention in addition to those shown anddescribed herein will be apparent to those skilled in the art from theforegoing description. Such modifications are also intended to fallwithin the scope of the appended claims.

What is claimed is:
 1. A polyphosphorylated compound for nuclearmagnetic resonance imaging of the gastrointestinal region whichcomprises a polyphosphorylated trimer, oligomer or polymer of analicyclic compound that is not a saccharide, said alicyclic compoundcomprising at least five carbon atoms, wherein said carbon atoms of saidpolyphosphorylated compound collectively contain at least two phosphatesubstituents and wherein all of said phosphate substituents are pendantfrom said carbon atoms of said polyphosphorylated compound for bindingto paramagnetic ions.
 2. A polyphosphorylated compound according toclaim 1 which is a polyphosphorylated polymer of alicyclic compound. 3.A polyphosphorylated compound according to claim 1 which is apolyphosphorylated trimer of a alicyclic compound.
 4. Apolyphosphorylated compound according to claim 1 which is apolyphosphorylated oligomer of alicyclic compound.
 5. Apolyphosphorylated compound according to claim 1 further comprising aparamagnetic ion.
 6. A polyphosphorylated compound according to claim 5wherein said paramagnetic ion is selected from the group consisting ofGd(III), Mn(II), Cu(II), Cr(III), Fe(II), Fe(III), Co(II), Er(II),Ni(II), Eu(III), Yb(III), and Dy(III).
 7. A polyphosphorylated compoundaccording to claim 6 wherein said paramagnetic ion is selected from thegroup consisting of Fe(III) and Mn(II).
 8. A polyphosphorylated compoundaccording to claim 7 wherein said paramagnetic ion is Mn(II).
 9. Apolyphosphorylated compound according to claim 1 further comprising apolyphosphorylated alicyclic compound having exactly one carbon ringstructure.
 10. A polyphosphorylated compound according to claim 1further comprising polyphosphorylated inositol.